Measurement of phosphatidylserine exposure during storage of platelet concentrates using the novel probe lactadherin: a comparison study with annexin V.
BACKGROUND
Annexin V binding to platelets (PLTs) is considered the gold standard for monitoring phosphatidylserine (PS) exposure. However, recent comparison of annexin V with the new calcium-independent PS probe lactadherin revealed that annexin V requires a certain threshold of PS exposure (2%-8%) for binding to occur. The aim of this study was to compare annexin V and lactadherin labeling of PLTs in PLT concentrates (PCs).


METHODS
Optimal labeling conditions for lactadherin and annexin V were established and then compared in either resting or calcium ionophore (CI)-activated PLTs from normal whole blood. Furthermore, 40 PCs (20 apheresis-derived and 20 pooled buffy coat-derived) were stored under standard blood bank conditions and PLT activation was monitored by measuring PS exposure with annexin V and lactadherin along with CD42b, CD61, and CD62P by flow cytometry on Days 1, 3, 5, and 7.


RESULTS
Lactadherin reported a higher exposure of PS than did annexin V in normal PLTs at submaximal doses of CI. PLTs from both types of concentrate, as expected, demonstrated evidence of increased activation during storage using annexin V, lactadherin, CD42b, or CD62P. However, a significantly higher percentage of PS-positive PLTs was found with lactadherin than annexin V.


CONCLUSIONS
PS exposure on the surface of stored PLTs has been previously underestimated due to the wide use of annexin V. Lactadherin provides a truer reflection of the degree of PS exposure and offers a new calcium-independent approach to studying PLT activation and/or apoptosis.
Albanyan
Abdul-Majeed
AM
Nuffield Department of Clinical Medicine, University of Oxford, Oxford, UK. abdul.albanyan@ndm.ox.ac.uk
Murphy
Michael F
MF
Rasmussen
Jan T
JT
Heegaard
Christian W
CW
Harrison
Paul
P
D017304:Annexin A5 / Q000737:chemistry* / Q000378:metabolism; D000954:Antigens, Surface / Q000737:chemistry* / Q000378:metabolism; D001792:Blood Platelets / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D039661:Integrin beta3 / Q000378:metabolism; D008894:Milk Proteins / Q000737:chemistry* / Q000378:metabolism; D015335:Molecular Probes / Q000737:chemistry* / Q000378:metabolism; D019007:P-Selectin / Q000378:metabolism; D010718:Phosphatidylserines / Q000378:metabolism* / Q000494:pharmacology; D015539:Platelet Activation / Q000187:drug effects; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism; D011309:Preservation, Biological; D011485:Protein Binding / Q000187:drug effects
D017304:Annexin A5; D000954:Antigens, Surface; C061248:ITGB3 protein, human; D039661:Integrin beta3; C098942:MFGE8 protein, human; D008894:Milk Proteins; D015335:Molecular Probes; D019007:P-Selectin; D010718:Phosphatidylserines; D019038:Platelet Glycoprotein GPIb-IX Complex